PROJECT SUMMARY/ABSTRACT? PROJECT 2 Project 2's objective is to understand the role for circular RNAs (circRNAs) in the mechanisms by which cocaine remodels the dorsal striatum (DS) and nucleus accumbens (NAc) to precipitate compulsive cocaine- seeking behaviors. circRNAs are highly conserved single-stranded ?back-spliced? RNAs in which the 5? and 3? ends of the transcript are covalently joined. Emerging evidence suggests that circRNAs are a major class of regulatory noncoding RNAs that are enriched in brain and that play key roles in basic aspects of neuronal function, but their involvement in the molecular, cellular, and behavioral actions of cocaine (and other drugs of abuse) has yet not been investigated. We will characterize patterns of circRNA expression in DS and NAc of mice that show compulsive-like cocaine consumption using paired-end ribominus RNA-sequencing. We will also assess circRNA expression in postmortem striatal tissues from humans with cocaine use disorders. We already have robust evidence for prominent cocaine regulation of several circRNAs in these brain regions. Those cocaine-responsive circRNAs that show similar abnormal expression in mice and humans will be prioritized for further investigation. We will determine whether prioritized cocaine-responsive circRNAs are regulated specifically in different types of DS and NAc neurons. We will then investigate the role played by these prioritized cocaine-responsive circRNAs in regulating the molecular and cellular responses to cocaine within these cell types of DS and NAc. This will be accomplished by use of in vivo CRISPR technology or RNAi to knockdown prioritized circRNAs in a cell type-specific manner and assess the influence of these circrRNAs in controlling baseline and cocaine-induced alterations in the intrinsic excitability of the neurons as well as their transcriptional responses to cocaine. Finally, we will investigate the ways in which cocaine-responsive circRNAs control compulsive-like cocaine self-administration behavior including relapse in mice. These highly innovative studies promise to yield fundamentally new insights into the molecular and cellular mechanisms of cocaine addiction, with parallel future studies planned for opiate drugs of abuse.